Details of a Researcher - ITAGAKI Shiro

Papers - ITAGAKI Shiro

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Exploration of candidate odorants derived from patient skin lesions using gas chromatography-mass spectrometry

Ryosuke IIZAWA, Masami HORIGUCHI, Hisashi UHARA, Yuji KAN, Kunihiro TOGASHI, Makoto SUZUKI, Takafumi NOMURA, Shiro ITAGAKI, Tomoko UNO, Madoka NAKAMURA

Journal of Japan Association on Odor Environment ( Japan Association on Odor Environment ) 55 ( 5 ) 315 - 318 2024.09 [Refereed]

DOI

A simple, dual direct expression plasmid system in prokaryotic and mammalian cells.

Manabu Murakami, Agnieszka M Murakami, Manabu Yonekura, Ichiro Miyoshi, Shirou Itagaki, Yasutaka Niwa

PNAS nexus 2 ( 5 ) pgad139 2023.05 [International journal]

We introduce a simple, dual direct cloning plasmid system (pgMAX-II) for gene expression analysis in both prokaryotic (Escherichia coli) and mammalian cells. This system, which uses a prokaryotic expression unit adapted from the pgMAX system and a mammalian promoter, is effective for subcloning using the DNA topoisomerase II toxin CcdB. Given that molecular biological cloning systems broadly rely on E. coli for rapid growth, the proposed concept may have wide applicability beyond mammalian cells.

DOI PubMed

Rapid method for plasmid DNA recombination (Murakami-system).

Agnieszka M Murakami, Manabu Yonekura, Katsuhiro Nagatomo, Yasutaka Niwa, Shirou Itagaki, Manabu Murakami

MethodsX 10 102167 - 102167 2023 [International journal]

DNA recombination is a useful technology for cloning and subsequent functional analysis, while standard techniques for plasmid DNA recombination have remained unchanged. In the present study, we introduced rapid method for plasmid DNA recombination, which we named "Murakami-system", to complete the experiments in under 33 h. For this purpose, we selected the following: PCR amplification with 25 cycles and E. coli strain with rapid growth (incubation time of 6-8 h). In addition, we selected rapid plasmid DNA purification (mini-prep; ∼10 min) and rapid restriction enzyme incubation (20 min). This recombination system enabled rapid plasmid DNA recombination within 24-33 h, which could be useful in various fields. We also established a 1-day method for competent cell preparation. Our rapid recombination system allowed several sessions of plasmid DNA recombination to be performed every week, which improves the functional analysis of various genes.•"Rapid method for plasmid DNA recombination (Murakami-system).•E. coli strain with rapid growth (incubation time of 6-8 h).•Combination of rapid protocols (PCR, electrophoresis, DNA purification, ligation, and mini-prep) enabled plasmid DNA recombination within 24-33 h.

DOI PubMed

Enhanced β-adrenergic response in mice with dominant-negative expression of the PKD2L1 channel

Manabu Murakami, Agnieszka M. Murakami, Takayuki Nemoto, Takayoshi Ohba, Manabu Yonekura, Yuichi Toyama, Hirofumi Tomita, Yasushi Matsuzaki, Daisuke Sawamura, Kazuyoshi Hirota, Shirou Itagaki, Yujiro Asada, Ichiro Miyoshi

PLOS ONE ( Public Library of Science (PLoS) ) 17 ( 1 ) e0261668 - e0261668 2022

Polycystic kidney disease (PKD) is the most common genetic cause of kidney failure in humans. Among the various PKD-related molecules, PKD2L1 forms cation channels, but its physiological importance is obscure. In the present study, we established a transgenic mouse line by overexpressing the dominant-negative form of the mouse PKD2L1 gene (i.e., lacking the pore-forming domain). The resulting PKD2L1del-Tg mice exhibited supraventricular premature contraction, as well as enhanced sensitivity to β-adrenergic stimulation and unstable R-R intervals in electrocardiography. During spontaneous atrial contraction, PKD2L1del-Tg atria showed enhanced sensitivity to isoproterenol, norepinephrine, and epinephrine. Action potential recording revealed a shortened action potential duration in PKD2L1del-Tg atria in response to isoproterenol. These findings indicated increased adrenergic sensitivity in PKD2L1del-Tg mice, suggesting that PKD2L1 is involved in sympathetic regulation.

DOI

Attenuated β-adrenergic response in calcium/calmodulin-dependent protein kinase IV-knockout mice.

Manabu Murakami, Agnieszka M Murakami, Yasushi Matsuzaki, Daisuke Sawamura, Takayoshi Ohba, Ichirou Miyoshi, Shirou Itagaki, Hiroyuki Sakagami

PloS one 16 ( 4 ) e0249932 2021 [Refereed] [International journal]

In the present study, we examined the importance of Ca2+/calmodulin-dependent protein kinase IV (CaMKIV) in the regulation of cardiac function using genetically modified CaMKIV-null mice. RT-PCR analysis revealed decreased expression of voltage-dependent calcium channels in the cardiac myocytes of CaMKIV-null mice compared with wild-type mice. CaMKIV-null mice showed shortened QT time on electrocardiograms. Pharmacological analysis revealed decreased responsiveness to the β-adrenergic blocker propranolol in CaMKIV-null mice, whereas the plasma norepinephrine level was not affected. CaMKIV-null mice showed decreased baroreflex on electrocardiograms. Heart rate variability analysis showed unstable R-R intervals, a decreased low frequency power/high frequency power (LF/HF) ratio, and increased standard deviation of the normal to normal R-R intervals (SDNN) in CaMKIV-null mice, suggesting decreased responsiveness to β-adrenergic stimulation in CaMKIV-null mice. Atrial contraction analysis and cardiac action potential recording showed a decreased response to the β-adrenoceptor agonist isoproterenol in CaMKIV-null mice. Furthermore, fluorescence imaging in a CRE-hrGFP assay revealed a decreased response to isoproterenol in CaMKIV-null cardiac myocytes. Taken together, our data strongly suggest a significant effect of CaMKIV gene ablation on cardiac β-adrenergic signal transduction.

DOI PubMed

Establishment of cardiac action potential recording using a membrane potential indicator in the mouse sinoatrial node.

Toyama Y, Yonekura M, Arakawa Y, Higuchi S, Tomita H, Murakami AM, Itagaki S, Murakami M

Hirosaki Med J 70 163 - 171 2020 [Refereed]

Molecular cloning of medaka orexin and orexin receptor and decreased spontaneous movement in response to an orexin inhibitor.

Niisawa T, Naruta N, Murakami AM, Yonekura M, Hirota K, Itagaki S, Murakami M

Hirosaki Med. J. 71 55 - 64 2020 [Refereed]

Decreased cardiac pacemaking and attenuated β-adrenergic response in TRIC-A knockout mice.

Manabu Murakami, Yuichi Toyama, Manabu Yonekura, Takayoshi Ohba, Yasushi Matsuzaki, Daisuke Sawamura, Agnieszka M Murakami, Miyuki Nishi, Shirou Itagaki, Hirofumi Tomita, Hiroshi Takeshima

PloS one 15 ( 12 ) e0244254 2020 [Refereed] [International journal]

Changes in intracellular calcium levels in the sinus node modulate cardiac pacemaking (the calcium clock). Trimeric intracellular cation (TRIC) channels are counterion channels on the surface of the sarcoplasmic reticulum and compensate for calcium release from ryanodine receptors, which play a major role in calcium-induced calcium release (CICR) and the calcium clock. TRIC channels are expected to affect the calcium clock in the sinus node. However, their physiological importance in cardiac rhythm formation remains unclear. We evaluated the importance of TRIC channels on cardiac pacemaking using TRIC-A-null (TRIC-A-/-) as well as TRIC-B+/-mice. Although systolic blood pressure (SBP) was not significantly different between wild-type (WT), TRIC-B+/-, and TRIC-A-/-mice, heart rate (HR) was significantly lower in TRIC-A-/-mice than other lines. Interestingly, HR and SBP showed a positive correlation in WT and TRIC-B+/-mice, while no such correlation was observed in TRIC-A-/-mice, suggesting modification of the blood pressure regulatory system in these mice. Isoproterenol (0.3 mg/kg) increased the HR in WT mice (98.8 ± 15.1 bpm), whereas a decreased response in HR was observed in TRIC-A-/-mice (23.8 ± 5.8 bpm), suggesting decreased sympathetic responses in TRIC-A-/-mice. Electrocardiography revealed unstable R-R intervals in TRIC-A-/-mice. Furthermore, TRIC-A-/-mice sometimes showed sinus pauses, suggesting a significant role of TRIC-A channels in cardiac pacemaking. In isolated atrium contraction or action potential recording, TRIC-A-/-mice showed decreased response to a β-adrenergic sympathetic nerve agonist (isoproterenol, 100 nM), indicating decreased sympathetic responses. In summary, TRIC-A-/-mice showed decreased cardiac pacemaking in the sinus node and attenuated responses to β-adrenergic stimulation, indicating the involvement of TRIC-A channels in cardiac rhythm formation and decreased sympathetic responses.

DOI PubMed

A simple and dual expression plasmid system in prokaryotic (E. coli) and mammalian cells.

Murakami M, Ohba T, Murakami A.M, Han C, Yonekura M, Kuwasako K, Itagaki S

Plos One, 14 e0216169 2019.05 [Refereed]

入院患者の健康食品・サプリメントと医薬品の相互作用リスクに関する実態調査

金森未侑, 冨澤登志子, 板垣史郎

日本老年薬学会雑誌 2 9 - 18 2019.04 [Refereed]

Medaka as a model for ECG analysis and the effect of verapamil

Manabu Yonekura, Nami Kondoh, Chong Han, Yuichi Toyama, Takayoshi Ohba, Kyoichi Ono, Shirou Itagaki, Hirofumi Tomita, Manabu Murakami

Journal of Pharmacological Sciences ( Japanese Pharmacological Society ) 137 ( 1 ) 55 - 60 2018.05 [Refereed]

The heart of the medaka, a small fish native to East Asia, has electrophysiological aspects similar to mammalian hearts. We found that the heart rates of medaka were more similar to humans than mice or rats. Medaka exhibited similar electrocardiogram patterns to those of humans, suggesting a similarity in cardiac impulse formation and propagation. Their hearts also exhibited similar responsiveness to verapamil, a calcium channel antagonist atropine, a parasympathetic nerve blocker propranolol, a sympathetic β-adrenergic blocker and isoproterenol, a sympathetic β-adrenergic agonist. We successfully analyzed action potentials and cardiac contractile forces in vivo. Verapamil affected action potential duration and reduced heart rate, suggesting the importance of voltage-dependent calcium channels in determining the heart rhythm of medaka. We also analyzed the expression of the voltage-dependent calcium channel β2 subunit, which participates in channel formation in cardiac myocytes, and found that splice variant type-2 was the only major transcript in the heart. Our results indicate that medaka could be an appropriate animal model for studying cardiovascular pharmacology.

DOI

Modified parasympathetic nervous system in Goto-Kakizaki diabetic rats.

Han C, Inaba W, Yonekura M, Ogata Y, Ohba T, Kushikata T, Niwa H, Yanagisawa T, Tomita H, Itagaki S, Hirota K, Akai H, Mizukami H, Murakami M

Hirosaki Med. J., ( 弘前大学大学院医学研究科・弘前医学会 ) 68 ( 1 ) 33 - 43 2017.10 [Refereed]

CiNii

Storage stability of serum formulations containing ofloxacin for autologous serum eardrop therapy

N. Hashimoto, K. Togami, K. Encou, H. Onodera, H. Tanimoto, S. Itagaki, S. Chono

PHARMAZIE ( GOVI-VERLAG PHARMAZEUTISCHER VERLAG GMBH ) 72 ( 3 ) 139 - 142 2017.03 [Refereed]

Objective: The storage stability of serum formulations containing ofloxacin for autologous serum eardrop therapy was evaluated for microbiological quality and component stability. Methods: Sterile serum formulations were prepared by mixing human serum and ofloxacin otic solution (1:1, v/v). To simulate eardrop contamination with external ear surface substances, prepared serum formulations were contaminated with a cotton swab that was rubbed sufficiently on the human external ear. Formulations were stored at 4 degrees C or room temperature in the dark. Colony forming units (CFUs), ofloxacin, and basic fibroblast growth factor (bFGF) concentrations in the stored serum formulations were determined. Results: The growth of microorganisms derived from the external ear was not detected in serum formulations after storage for 14 days, regardless of temperature. However, microbial growth was detected in serum formulations stored without ofloxacin, indicating that this is necessary for storage. In addition, concentrations of ofloxacin and bFGF did not decrease over 14 days, indicating that ofloxacin and bFGF in serum formulations are stable for this time period. Conclusion: The present study indicates that the efficacy and safety of serum formulations used as a therapy for perforated eardrums are stable and safe for at least 14 days.

DOI

Effects of halothane anesthesia on electrocardiogram parameters in mice.

Nozaki Shuhei, Ogata Yoshiki, Yonekura Manabu, Han Chong, Niwa Hidetoshi, Kushikata Tetsuya, Hirota Kazuyoshi, Matsuzaki Yasushi, Tomita Hirofumi, Imaizumi Tadaatsu, Itagaki Shirou, Sawamura Daisuke, Murakami Manabu

Hirosaki Med. J. ( 弘前大学大学院医学研究科・弘前医学会 ) 67 ( 2 ) 129 - 135 2017.02 [Refereed]

We analyzed the effects of halothane as an inhalational anesthetic agent on electrocardiogram （ECG）parameters in C57/BL6 mice. Following induction of anesthesia using 2% halothane, the ECGs showed a regularpattern and the heart rate （HR） was within an acceptable range （~500 bpm）. The HR decreased with increasinghalothane concentration in a concentration-dependent fashion. The frequency domain analysis showed that the high-frequency （HF） component decreased and the lowfrequency（LF） component increased in a concentration-dependent fashion. Therefore, the LF/HF ratio increasedwith increasing halothane concentration, suggesting effects on the autonomic nervous system. We analyzed the pharmacological response to sympathetic blockade with propranolol, a typical adrenergicβ-blocker, under halothane anesthesia. Propranolol administration resulted in a decreased HR; interestingly,intraperitoneal injection of propranolol （120 μg/kg body weight） resulted in arrhythmia （sick sinus syndrome）during anesthesia with 3% halothane. Our results indicate the importance of selecting a suitable anesthetic agent for C57/BL6 mice in pharmacological studies

CiNii

Aerosolized liposomes with dipalmitoyl phosphatidylcholine enhance pulmonary absorption of encapsulated insulin compared with co-administered insulin

Sumio Chono, Kohei Togami, Shirou Itagaki

DRUG DEVELOPMENT AND INDUSTRIAL PHARMACY ( TAYLOR & FRANCIS LTD ) 43 ( 11 ) 1892 - 1898 2017 [Refereed]

Objective: We have previously shown that aerosolized liposomes with dipalmitoyl phosphatidylcholine (DPPC) enhance the pulmonary absorption of encapsulated insulin. In this study, we aimed to compare insulin encapsulated into the liposomes versus co-administration of empty liposomes and unencapsulated free insulin, where the DPCC liposomes would serve as absorption enhancer. Significance: The present study provides the useful information for development of noninvasive treatment of diabetes. Methods: Co-administration of empty DPPC liposomes and unencapsulated free insulin was investigated in vivo to assess the potential enhancement in protein pulmonary absorption. Co-administration was compared to DPPC liposomes encapsulating insulin, and free insulin. Results: DPPC liposomes enhanced the pulmonary absorption of unencapsulated free insulin; however, the enhancing effect was lower than that of the DPPC liposomes encapsulating insulin. The mechanism of the pulmonary absorption of unencapsulated free insulin by DPPC liposomes involved the opening of epithelial cell space in alveolar mucosa, and not mucosal cell damage, similar to that of the DPPC liposomes encapsulating insulin. In an in vitro stability test, insulin in the alveolar mucus layer that covers epithelial cells was stable. These findings suggest that, although unencapsulated free insulin spreads throughout the alveolar mucus layer, the concentration of insulin released near the absorption surface is increased by the encapsulation of insulin into DPPC liposomes and the absorption efficiency is also increased. Conclusion: We revealed that the encapsulation of insulin into DPPC liposomes is more effective for pulmonary insulin absorption than co-administration of DPPC liposomes and unencapsulated free insulin.

DOI PubMed

Sustained distribution of aerosolized PEGylated liposomes in epithelial lining fluids on alveolar surfaces

Keita Kaneko, Kohei Togami, Eri Yamamoto, Shujun Wang, Kazuhiro Morimoto, Shirou Itagaki, Sumio Chono

DRUG DELIVERY AND TRANSLATIONAL RESEARCH ( SPRINGER HEIDELBERG ) 6 ( 5 ) 565 - 571 2016.10 [Refereed]

The distribution characteristics of aerosolized PEGylated liposomes in alveolar epithelial lining fluid (ELF) were examined in rats, and the ensuing mechanisms were investigated in the in vitro uptake and protein adsorption experiments. Nonmodified or PEGylated liposomes (particle size 100 nm) were aerosolized into rat lungs. PEGylated liposomes were distributed more sustainably in ELFs than nonmodified liposomes. Furthermore, the uptake of PEGylated liposomes by alveolar macrophages (AMs) was less than that of nonmodified liposomes. In further in vitro uptake experiments, nonmodified and PEGylated liposomes were opsonized with rat ELF components and then added to NR8383 cells as cultured rat AMs. The uptake of opsonized PEGylated liposomes by NR8383 cells was lower than that of opsonized nonmodified liposomes. Moreover, the protein absorption levels in opsonized PEGylated liposomes were lower than those in opsonized nonmodified liposomes. These findings suggest that sustained distributions of aerosolized PEGylated liposomes in ELFs reflect evasion of liposomal opsonization with surfactant proteins and consequent reductions in uptake by AMs. These data indicate the potential of PEGylated liposomes as aerosol-based drug delivery system that target ELF for the treatment of respiratory diseases.

DOI PubMed

「感染性心内膜炎薬物治療の手引き」を用いた抗菌薬適正使用への貢献

小笠原真帆, 太田, 岡村祐嗣, 津山博匡, 板垣史郎, 萱場広之, 早狩誠

日本病院薬剤師会雑誌 ( (一社)日本病院薬剤師会 ) 52 ( 10 ) 1287 - 1292 2016.10 [Refereed]

感染防止対策加算連携施設間における抗菌薬使用量サーベイランスの評価

岡村祐嗣, 倉内寿孝, 津山博匡, 板垣史郎, 萱場広之, 早狩誠

日本環境感染学会誌 ( (一社)日本環境感染学会 ) 31 ( 5 ) 326 - 334 2016.09 [Refereed]

The present study used antimicrobial consumption surveillance to assess the resistance rate transition of Pseudomonas aeruginosa in facilities cooperating in infection prevention medical fees using the antimicrobial use density (AUD) and days of therapy (DOT) as parameters, and compared the surveillance results in 2014 to 2012. In our hospital, the change in DOT of first-generation cephalosporins, third-generation cephalosporins, fourth-generation cephalosporins, carbapenems, aminoglycosides, glycopeptides was greater than that of AUD. In stroke center, the change in DOT of penicillins, antipseudomonal penicillins, carbapenems was greater than that of AUD. The surveillance comparing the AUD transition with DOT transition revealed the influence of antimicrobial dose and duration on the change of AUD was more accurately evaluated by surveillance using the AUD transition with DOT transition. In both facilities, the antimicrobial dose and duration were not associated with change in the resistance rate of P. aeruginosa. The present results suggest that the effects of change in antimicrobial dose and duration transition on the change in resistance rate of P. aeruginosa are limited.

DOI CiNii

ヒト官能試験による口腔内崩壊錠の服用性の実証

丁野純男, 谷本裕幸, 松井萌, 中村勝貴, 佐藤卓馬, 中川麻奈, 山田恵乃, 金子圭太, 山佳織, 佐藤恵亮, 山下美妃, 丹保好子, 郡修徳, 森本一洋, 板垣史郎, 葛西亮也

臨床と研究 93 ( 5 ) 171 - 174 2016.05

膵臓がん組織の抗がん剤反応性を予測するバイオマーカーの探索

板垣史郎

医療の広場, 55 13 - 16 2015.04 [Invited]

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Papers - ITAGAKI Shiro